Disposable laboratory implement

ABSTRACT

A plastic, disposable laboratory implement used in treating and processing liquid samples containing nucleic acids. The implement includes at least one wall segment in contact with the liquid sample and made of a polypropylene mixed with at least one additive. The additive lowers a bonding affinity of the polypropylene for nucleic acids. The wall segment exhibits a surface gloss&gt;160 as measured per DIN 67530 at an angle of 60°.

The present invention relates to a single-use, hereafter disposable,plastic laboratory implement defined in the preamble of claim 1.

In particular disposable implements of this kind are reactionreceptacles, pipet tips and also microtitration plates. In general thepresent invention applies to all plastic implements used in thelaboratory that may be applicable in processing liquid samplescontaining nucleic acids.

Disposable laboratory implements of this kind are made of polypropylene.It has been widely observed that nucleic acids interact withconventional polypropylenes in a manner that under some conditions thenucleic acids will bond for instance to the walls of the reactionreceptacles.

The interaction between nucleic acids and different polypropylenereaction receptacles is described for instance in CLINICAL NOTES ofMarch 2001, pp 52. This article cites a fact also observed by theapplicant that conventional polypropylenes will bond nucleic acidsespecially at high salt concentrations. The processing of nucleic acidsentailing various steps in the phase transition range, that is at highsalt concentrations, the possibility of impoverishing the nucleic acidsof interest when using conventional polypropylene implements cannot beexcluded.

The publication above mentions that all tested conventional propylenereceptacles exhibit substantially the same adsorption properties asregards nucleic acids. A few receptacles made with special materialsexhibited less bonding for nucleic acids; however the source of thisfeature could not be ascertained for lack of manfacturer data.

U.S. Pat. No. 6,544,417 discloses making illustratively polypropylenelaboratory implements by adding additives to them in a manner that thebiomolecule's bonding ability shall be reduced. The additives describedin this U.S. Pat. No. 6,544,417 always are other plastics, for instancefluoropolymers such as teflon. Rigorously speaking, the compositionsdescribed therein are not plastics containing additives but blends orcompounds of two different plastics. Contrary to additive containingplastics, the mixtures of materials known from the above US patent donot permit making transparent laboratory implements.

The objective of the present invention is to create transparentpolypropylene laboratory implements and to manufacture them inespecially simple manner, said implements of the present inventionexhibiting in particular in the critical high salt range, a lowerbonding affinity for nucleic acids than do conventional laboratoryimplements.

This problem is solved by means of a laboratory implement defined by thefeatures of claim 1.

Applicant surprisingly discovered that polypropylene surfaces exhibitingsubstantially higher surface gloss than standard polyproplyenes offerthe lower bonding affinity to nucleic acids, especially under high saltconditions, which is sought in the present invention.

The expression “surface gloss” herein denotes the light reflectingproperty of surfaces. This surface gloss is defined as being theintensity of light reflected by a tested surface. Accordingly opticallymatte surfaces exhibit a value less than 10, medium glossy surfacesexhibit values between 10 and 70 and high-gloss surfaces values>70.Conventional polypropylenes exhibit a surface gloss of about 90.

The gloss values of the particular surface as a rule are measured atdifferent angles of incidence (for instance 20, 60 or 85°). Suchmeasurements are carried out in internationally uniform manner accordingto DIN 67530 (Publication date: 1982-01), ISO 2813 (1994/Cor. 1997) orASTM D523-89 (1999).

The values of the present invention all relate to measurements at 60°.The surface coefficient may be measured using commercially availableapparatus. Illustratively MELIT Co. offers their “PicoGloss 560” whichallows simple surface gloss measurements in the range which issignificant for polypropylene.

Applicant presumes that, compared with conventional polypropylenes,those exhibiting the surface gloss of the invention exhibit fewerinitial defects for instance in the form of edge roughness that, in theconventionally used polypropylenes may act as seed crystals and triggerthe bonding of the nucleic acids to the walls of the disposablelaboratory receptacles.

Polypropylenes exhibiting the surface gloss of the inventionsubstantially differ from the conventional polypropylenes in that theyshow considerably smaller crystalline polypropylene units in the surfacezones. This feature is attained using appropriate additives that willdissolve in the melt and upon solidification will precipitate as finelydistributed seed crystals. The smaller the polypropylene units (the morecrystallites available during solidification), the clearer thepolypropylene shall be. Especially appropriate additives are theso-called clarifiers. A clarifier marketed under the name ADK STAB NA-21for instance allows making polypropylenes exhibiting especially highsurface gloss.

As discussed above, at least one wall zone making contact with theliquid sample is made from an additive-containing polypropylene wheresaid additive introduces especially high gloss to a plastic surfacewhich then evinces the desired reduced bonding affinity to nucleic acidsat high salt conditions.

Within the scope of the invention basically all additives increasingpolypropylene's surface gloss are applicable, but in especiallypreferred manner the said additive clarifier ADK STAB Na-21 offered byADEKA PALMAROLA SAS (Strasbourg, France) will be used. This substance isaluminum hydroxybis[2,2′-methylenebis (4,6-itert.butylphenyl)phosphate].Obviously other clarifiers and other additives allowing adjusting thesurface gloss of the invention, such as the products MILLAD 3988, MILLAD3950 and HPN-68 made by MILLIKAN Corp. or NC-4 made by MITSUI TOATSU Co.also are applicable. These products are diverse clarifiers generatingthe above discussed fine surface structure on the molded polypropylenepart. Moreover additives such as polypropylene waxes may also be used toincrease the surface gloss of polypropylenes.

To-date clarifiers have been added to plastics in the state of the artfor the purpose of increasing implement transparency. The conventionalconcentrations of the above clarifier ADK Stab NA-21 are 0.09% (wt/wt)referred to the total weight. At such concentrations a slightly reducedbonding affinity of the polypropylene mixed with the clarifier alreadymay be observed. However this slight effect is quite insufficient toprocess samples containing nucleic acids absent significant losses. Thedefinite reduction in bonding affinity attained by the invention betweenpolypropylene implements and nucleic acids will be sensible only whenADK Stab NA-21 is added to polypropylene in concentrations above 0.2%(wt/wt). Tests run by applicant show a minimum of 0.4% (wt/wt) of thesaid clarifier must be added to the polypropylene implements to keep theloss of nucleic acids during processing with the polypropyleneimplements at <1 (wt/wt) [relative to the initial quantity of nucleicacids]. Thus the clarifier concentrations selected in the invention toattain the desired bonding properties is much above the concentrationswhich are conventionally required for transparent plastic implements,the applicant having been first in discovering that, surprisingly, whenadding unusually high clarifier concentrations, there results a dramaticchange in the bonding behavior of polypropylene as compared to knowntransparent plastic implements.

The laboratory implements preferably may be pipet tips, syringes, vialsstoring liquid samples, microtitration plates, further bioarray slides,pestles or agitators etc. However the invention is not restricted tothese implements. In principle those implements also are covered which,within the scope of processing liquid samples containing nucleic acids,will be in contact with the samples over an extended time interval.

According to the invention, at least the wall segments of the disposablelaboratory implement that make contact with the liquid sample shall bemade of a polypropylene mixed with an additive. Illustratively and inparticular as regards microtitration plates, only the reactionreceptacles (wells) need be made of polypropylene exhibiting the surfacegloss of the invention whereas a frame supporting the reactionreceptacles consists of another plastic, for instance a polycarbonate.

On the other hand, as regards other, more economical disposablelaboratory implements, and in a preferred embodiment mode of theinvention, the implements all are made from the additive-mixedpolypropylene and as result all of them exhibit the surface gloss of theinvention.

The invention is elucidated below in relation to several embodimentmodes.

1. Measuring DNA Absorption as a Function of Gloss Coefficient.

Micro-reaction receptacles made by injection-molding polypropylenesfitted with various additives were used to measure the DNA adsorption atpolypropylene surfaces by filling them with 50 μltr of a radioactivelymarked DNA solution (0.2 ng of DNA/μltr) at a 2.5 molar NaClconcentration and storing them in one test preparation for 24 h at 37°C. and in another test preparation for 10 min at 95° C. Then thesolution was pipetted, that is completely evacuated. Next the reactionreceptacle emptied in this manner was checked for its residualradioactivity. In this manner the DNA portion that was lost byadsorption in the reaction receptacle could be determinedquantitatively.

It was found that reaction receptacles made of propylene to which theclarifiers ADK Stab NA-21, MILLAD 3988 and MILLAD 3950 exhibitedconsiderably reduced DNA adsorption at the receptacle walls. Because theabove additives increase the gloss of molded propylene surfaces—asdetermined on test bodies by the manufacturers of additives—the DNAadsorptivities may be correlated to the surface gloss and moreover amodel may be developed (see above) to explain the adsorptiondifferentials. The Table below shows numerical correlation values. TABLEDNA adsorption and gloss coefficient of polypropylenes with differentadditive treatments of polypropylene receptacles DNA adsorption DNAadsorption Gloss (37° C., 24 h) (95° C., 10 min) coefficient Material %wt/wt) (wt/wt) at 60° [−]* polypropylene 65-95 >90 90 no clarifierpolypropylene + 2.5 3.0 165 0.3% (wt/wt) MILLAD 3950 polypropylene + 1.12.01 165 0.3% (wt/wt) MILLAD 3988 polypropylene + 0.7 1.0 175 0.3%(wt/wt) ADK Stab NA-21(*from Adeka Palmarole Deutschland GmbH)2. Manufacturing a Disposable Laboratory Implement

The polypropylene receptacles are made by standard injection molding ofpolypropylene granulates. The corresponding additive, i.e. clarifier iseither admixed using a master batch (a concentrate of the additive inpolypropylene) as a granulate to the basic polypropylene granulate (thatis, the two granulates are physically mixed as a dry blend and themixture of granulates then is injection molded) or the additive shallalready be contained from the beginning in the basic polypropylenegranulate and is delivered from the manufacturer as the finishedproduct. Furthermore the additive in the form of a pure substance may beadmixed by using a compounding unit, for instance using a twin wormextruder, at the final desired proportion, into the polypropylene meltand to granulate the material from the melt after solidification. Thisgranulate may then be injection molded.

1. A plastic, disposable laboratory implement used in treating andprocessing liquid samples containing nucleic acids, said implementcomprising at least one wall segment in contact with the liquid sampleand being made of a polypropylene mixed with at least one additive, saidadditive lowering a bonding affinity of the polypropylene for nucleicacids, wherein the wall segment exhibits a surface gloss>160 as measuredper DIN 67530 at an angle of 60°.
 2. The disposable laboratory implementas claimed in claim 1, wherein the wall segment exhibits a surfacegloss>170.
 3. The disposable laboratory implement as claimed in claim 1,wherein the additive is a clarifier.
 4. The disposable laboratoryimplement as claimed in claim 3, wherein the clarifier is contained at aconcentration in the polypropylene to that exceeds concentrationsconventionally used with transparent implements and said clarifierconcentration being in a range of concentrations wherein increasing theclarifier concentration no longer results in a significant increase intransparency.
 5. The disposable laboratory implement as claimed in claim3, wherein the clarifier is an available substance designated as ADKSTAB NA-21.
 6. The disposable laboratory implement as claimed in claim5, wherein the clarifier ADK Stab NA-21 is contained in aconcentration>0.2% (wt/wt) in the polypropylene.
 7. The disposablelaboratory implement as claimed in claim 1, wherein said implementserves as a device selected from the group consisting of pipet tips,reaction receptacles, receptacles storing liquid samples andmicrotitration plates.
 8. The disposable laboratory implement as claimedin claim 1, wherein the disposable laboratory implement is integrallymade of the polypropylene mixed with at least one additive.
 9. Thedisposable laboratory implement as claimed in claim 2, wherein theadditive is a clarifier.
 10. The disposable laboratory implement asclaimed in claim 9, wherein the clarifier is contained at aconcentration in the polypropylene that exceeds concentrationsconventionally used with transparent implements and said clarifierconcentration being in a range of concentrations wherein increasing theclarifier concentration no longer results in a significant increase intransparency.
 11. The disposable laboratory implement as claimed inclaim 9, wherein the clarifier is an available substance designated asADK STAB NA-21.
 12. The disposable laboratory implement as claimed inclaim 10, wherein the clarifier is an available substance designated asADK STAB NA-21.
 13. The disposable laboratory implement as claimed inclaim 11, wherein the clarifier ADK Stab NA-21 is contained in aconcentration>0.2% (wt/wt) in the polypropylene.
 14. The disposablelaboratory implement as claimed in claim 12, wherein the clarifier ADKStab NA-21 is contained in a concentration>0.2% (wt/wt) in thepolypropylene.